Our ability to answer scientific questions is always constrained by the technology available. With the advent of new technologies, answers that were once out of reach become attainable. Therefore an important function of our Centre is to continually develop and apply the newest technologies, and where possible, make them available for other researchers across Australia (for example via the Australian Plant Phenomics Facility and Bioplatforms Australia).
The following Centre technology platforms are used to answer our scientific questions:
Arabidopsis phenotyping platform
Measuring plant growth and function (phenotyping) is key to identifying the roles of the genes, proteins and regulators that we study. Our range of controlled growth cabinets (> 100 m ) and glasshouse space (>60 m ) allow measurement of plant growth under varying light, temperature and CO2 conditions across all 3 nodes. Our phenomic analysis platforms include:
- Gas exchange systems for analysis of CO and O2 exchange, including isotope analysis to monitor photosynthesis and respiration. In 2012 we installed several new plant growth rooms and purchased a multiplexed gas exchange system from Qubit Systems, funded by ARC LIEF.
- Chlorophyll fluorescence imaging systems to monitor spatial and temporal changes in leaf chloroplast properties both in high resolution and high throughput.
- Imaging-based growth analysis systems for monitoring the growth of plant shoots under various conditions. In 2012, ARC LIEF funded a joint project between UWA and ANU Centre researchers to build new climatemimicking growth environments with sophisticated LED lighting and cameras to build climate scenarios and measure plant responses. Our in-house facilities are complemented by the NCRIS-funded Australian Plant Phenomics Facility, with CI's Pogson and Badger on the Executive Management Committee of the High-Resolution Plant Phenotyping Centre in Canberra.
- An extracellular flux analyser from Seahorse Biosciences, being used for the first time in plants to measure respiratory and glycolytic rates using fluorescence ion selective probes. In addition, our new Oroboros Oxygraph-2K for simultaneous measurements of oxygen consumption rate and reactive oxygen species production has applications for a variety of centre projects working on the role of ROS as a signalling molecule in plants.
In addition to our in-house facilities, the Centre is closely involved with the NCRIS-funded Australian Plant Phenomics Facility, and particularly with the High-Resolution Plant Phenotyping Centre (HRPPC) based in Canberra.
Molecular profiling platforms
The first steps in molecular profiling involve isolating the cells to be profiled and extracting the molecules that we wish to identify and quantify. Increasingly we need to be able to deal with very small samples and high numbers of samples in parallel. Two new platforms relevant to these needs were acquired in 2012.
Firstly, we obtained a laser capture micro-dissection platform (Zeiss PALM MicroBeam). Precision lasers allow the capture of cells and cell components from fresh, paraffinized or frozen plant tissue sections to create accurate and repeatable molecular analysis of DNA, RNA and proteins.
Secondly, our new platform at ANU will include a robot for high-throughput sample preparation from frozen plant tissue, funded from ARC LIEF. This will accelerate our extractions of DNA and metabolites from large numbers of samples, making large-scale projects feasible that weren't previously.
Once we have taken these samples from the plants of interest, the following technology allows us to analyse the DNA, RNA, proteins or metabolites.
Both UWA and ANU have next-generation deep sequencing platforms installed. Our Illumina Hi-Seq Deep Sequencer is one of the most powerful platforms for next generation sequencing. This platform can analyse both DNA and RNA samples, i.e. genome and transcriptome sequencing.
“In a single day of use, this deep sequencing technology will allow researchers to obtain the sequence equivalent of the entire human genome project, which took 4 billion dollars and 10 years to complete over a decade ago”
Our transcriptomics platform consists of a complete Affymetrix microarray platform, Roche 480 Lightcyclers for high- throughput real-time PCR, and associated support instruments such as Bioanalysers and robotics for liquid handling. Future transcriptomics projects will benefit from the deep-sequencing platform.
The principal tools we use in the Centre for proteome analysis are electrophoresis are our four mass spectrometers:
These mass spectrometers enable us to identify novel proteins, as well as determine quite small changes in protein quantities, generated either by changes in levels of gene transcription, translation of mRNA, protein degradation, or post-translational modification. We collaborate closely with Agilent Technologies and Bruker Biosciences to develop new protocols for the application of this cutting-edge technology.
Our principal tools in metabolomics are separation technologies (organelle separation by centrifugation; metabolite separation by liquid and gas chromatography) and mass spectrometry. Separated molecules are fed into a mass spectrometer that fragments them and identifies them according to the spectrum of fragment masses. We obtain quantitative data for potentially hundreds of energy metabolites in a single mass spectrometric analysis.
We have also taken an active role in developing a metabolomics database and analysis software which is available to researchers in the Centre and at International institutions with our Metabolome Express website (www.metabolome-express.org).
We were key proponents of the $9.5M NCRIS investment in Metabolomics Australia and play an important management role in the facilities installed in WA. The Centre thus has access to some of most modern metabolomics facilities anywhere in Australia.
Membrane Transporter Expression Facility
Communication and energy exchange between organelles and between cells requires highly specialised protein channels and transporters. To fully understand their function, they must be expressed in cells where they can be characterized in isolation from other related transport proteins. This can be done in a cell where there is either a low background of native transport (Xenopus laevis oocytes) or in cells where particular transport proteins are silenced (usually yeast). Such heterologous expression is essential for elucidating function, identifying substrate specificity, and investigating post-translational regulatory processes. The UA node houses the facility, originally set-up by a LIEF grant and funding from the SA government.
Our informatics expertise and resources benefit greatly from the Centre of Excellence in Computational Systems Biology funded through the WA Centres of Excellence in Science and Innovation scheme. The Computational Systems Biology centre is working closely with the ARC Centre of Excellence in Plant Energy Biology.
The Centre is linked to the Western Australian Supercomputing Program (WASP), one of Australia's most prestigious and internationally competitive supercomputer and scientific visualisation centres. Through WASP, the Centre has access to the iVEC network of high-performance computing facilities across Perth.
The Centre has constructed several databases to house and analyse molecular profiling data integrated into genomic data sets based on Arabidopsis genome sequencing and annotation from international sources.
Web-based interfaces to our databases and software are available on the tools page of this website.